Antibody-mediated depletion is widely used as induction therapy in sensitized transplant recipients to overcome the deleterious effects of preexisting donor-reactive immunity. However, memory T cells are less susceptible to depletion than nave T cells and there are no current efforts to improve the efficacy of induction therapies in targeting donor-reactive T cell memory. Our previous studies in a mouse model of cardiac transplantation using rabbit anti-murine thymoglobulin (mATG) showed that recovering memory CD8 T cells are the primary effector mechanism mediating allograft rejection in mATG treated recipients. Preliminary experiments showed that help from depletion-resistant memory CD4 T cells is required for memory CD8 T cell recovery following antibody-mediated depletion and identified B cells as critical mediators of this help. Limiting CD4 T cell help delays memory CD8 T cell reconstitution markedly enhances the efficacy of mATG induction therapy in sensitized recipients and is associated with the increase in T and B cells with regulatory phenotypes. The goal of the proposed study is to determine the mechanisms of memory T cell reconstitution following antibody mediated depletion and to use this information to develop strategies improving allograft outcome in high risk recipients. We hypothesize that following antibody-mediated lymphoablation, residual memory CD4 T cells interact with B cells through TCR/MHC class II and CD40/CD154 and induce Beff cell activation to produce inflammatory cytokines such as TNF?. Subsequently, cognate B cell/CD8 T cell interactions and B cell derived cytokines facilitate homeostatic CD8 T cell expansion. Interference with helper signals will increase Breg/Beff and Treg/Teff cell ratios and effectively inhibit memory CD8 T cell recovery and the development of pathogenic anti-donor responses. We will test this hypothesis in three Specific Aims: Aim 1. To determine the mechanisms through which B cells mediate memory CD8 T cell reconstitution following antibody-mediated lymphoablation. Aim 2. To test whether blocking CD4 T cell help increases the efficacy of mATG induction therapy in sensitized heart allograft recipients. Aim 3. To test the contribution of regulatory T cells to allograft prolongatin by lymphoablative strategies. We anticipate that the approaches developed in these studies will specifically target pre-existing donor-reactive memory T cells and will improve the efficacy of lymphoablation in sensitized transplant patients.